I have an antibody solution at a concentration of 200 micrograms/ml, as supplied by the manufacturer. I stored it at 4 degree, as stated in the data sheet.
I will use this 200 micrograms for western blotting and immunohistochemistry. I am ready to perform the immuohistochemical study, however, I have to wait for a month or two till western blotting.
My question is if I opened the vial now and pipetting 30 micrograms of the antibody solution and restore it for a month or two, would this affect the antibody stability?. I am worried about introducing any microbial organism or air in the stock vial?
Do you recommend postponing the immunohistochemical study till I am ready for western blotting?
Well the storage of opened vial can be at -20 or -80 degrees if you have any doubts about that. Also contamination is not the problem if u use sterile tip's for getting 30ug that u sad. But longer than 2 weeks after opening its better to be frozen. Also if you are not in a hurry you can postpone your staining.
all best
Better to preserve the aliquotes it at -20 degree for long term storage. Avoid repeat freeze thaw cycles. If thawed once keep refrigerated at 4 degree up to consumption.
I agree with Rudra. For long term experiments, store small aliquots (just enough for a single experiment) in eppindorf tubes at -80C and thaw when required.
you definitely want to avoid multiple freeze/thaws of your antibody. As stated previously, you should aliquot your antibody so that you only have 1 thawing per use. You can store your antibody at -20 for a long period of time, but -80 is best for longer periods as stated by Edmund. If you are really concerned about stability of your antibody you could always lyophilize your antibody.
Store small aliquots at -20. However, repeated freezing and thawing of aliquots should be avoided, so try to make aliquots that would make sense for how much you use. Just enough for an experiment, without much left over. Once you thaw an aliquot, store at 4 degrees. Dating the thaw date helps make sure you are using it within a month or two. However, a while back I learned a more economical solution, and this is what I now do: mix your antibody with equal parts of glycerol (sterilize glycerol by autoclaving before hand). This 50% solution will not freeze, and can be taken in and out of -20 without freezing/thawing and damaging the protein structure of the antibody. Put it o/n on a rocker in the cold room to mix thoroughly, then store in the -20 in 2-3 larger aliquots. These can be taken in and out of the -20, and placed on ice in the interim. As stated above though, the best for long term storage is aliquots at -80.
thank you so much for your answers
however, my datasheet of the antibody stated " DO not freeze", and it also states that the antibody is stable for one year at 4 degrees.
I also contacted the technique service and they said that there s no need to aliquot the antibody.
do you still recommend after that to aliquot the antibody in -20 or -80?
if your antibody concentration is only 200 ug/mL you are most likely going to have to use a fairly high concentration for western blots. If the antibody is stable for 1 year at 4 degrees you might use the most of it over the course of the year so you would not have to worry about aliquoting it. I agree with karen's post that you can add glycerol to the antibody and put it in -20 and it will be more stable, yet not freeze. If you think that the antibody will last longer than 1 year with your uses, I would follow Karen's instructions. good luck
thank you so much again, for all your answers
you have been so helpful. I think you are all agree not to open the manufacture's tube till I am ready for both IHC and WB.
after that, adding 50% glycerol and aliquoting at -80 would be the best option to avoid freezing of the antibody.
I hope that my uses won't extend beyond 2 months from now, God willng
if I have any misunderstanding, please help.
Check if there is a preservative included in the solution. If not, add sodium azide to a final concentration of 0.1% m/v. (eg from a 10% m/v stock prepared in water or PBS).
- Badges
- Science topic
- Similar topics
- Biological Science
- Immunology
- Antibodies